AD-A143 082 EVALUATION INNRHESUS MONKEYS OF A BI VALENT yIEATTENUATED DENGUE VACCINE CONTAINING TYP ES 2 AND 4 AVIRUSES U) PUERT0 RICO UNIV SAN JUAN E NRAISEIBURD

UNCLASSIFIED JUN 84 DAMO 783-C 3029 F/G6/T

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00o "Evaluation in Rhesus Monkeys of a Bivalent Live

Attenuated Dengue Vaccine Containing Types 2 and4and 4 Virusess"

UAnnual and Final Report

Ediunco Kraiselburd, Ph.D.

June 1984

Supported by

Us Army Medical Research and Development CommandFort Detrick, Frederick, Maryland 21701

Contract No. DAMD7-83-C-3029

University of Puerto RicoMedical Sciences Carpus, GPO Box 5067San Juan, Puerto Rico 00936

-DTIC0.. EECTE

CD JUL 1 2 1984V

WLJ MD Distribution Statenent S DLi- Approved for public release; distribution unlimited D

The findings in this report are not to be construed asan official Depatent of the A=T position unles 80designated by other authorized domients.

84 07 10 089, ... . ' - == '.W-O N ,

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REPORT DOCMENTATION PAGE DEFOHE CONII1LETING FORMI. REPONT NUMBER 24~ . GOVT Aj( E~OO RCIPEN T5S CAT ALOG NUM#9Rf

4. TiTL.E t"k .. ,. S. TYPE OF REPORT A pEmoD COVEREDEvaluation in Rhesus Monkeys of a Bivalent LiveAttenuated Dengue Vaccine containing Types 2 and Final (Dec. 1/84 - lMy 31/844 Viruses. s. PERFORMING ORG. REPORT NUMBER

7." AUTHOR(* A. CONTRACT OR GRANT NUMBER(e)

Edrpido Kraiselburd, Ph.D. D17-83-C-3029

9. PERFORMING ORGANIZATION NAME AND ADDRESS 10. PROGRAM ELEMENT. PROJECT. TASK

University of Puerto Rico AREA & WORK UNIT NUMBERS

Medical Sciences CawpusSan Juan, Puerto Rico 00936

11. CONTROLLING OFFICE NAME AND ADDRESS 12. REPORT DATE

US Anuy Medical Research and Development Comnand' June 1984Fort Detrick 3. NUMBER OF PAGESFrederick, Maryland 21701 6

I4. MONITORING AGENCY NAME A ADDRESS(JI dierent Irm CentroJlne 0*1ae.) IS. SECURITY CLASS. (of tls rer.,t)

IS&. DECL ASSIFICATONDOWNGRADINGSCHEDULE

I6. DISTRIBUTION STATEMENT (of this Report)

Approvqd for public release, distribution unlimited

17. DISTRIBUTION STATEMENT (of &he 'birect ehered in Blec 20, it dlfferm Iram iep)

111 SUPPLEMENTARY NOTES

IS. KEY WORDS (Gm.enmue on owoe de it nbeoss ry md Identfly AW black numbef)

DMI-2/S-1 Vaccine; DM-4 Vaccine Rhesus monkeys imuunty.

The purpose of this study was to detenuine whether a bivalent (dengue-2-4)vaccine muld confer iurmity in rhesus mkeys against wild type virus infection

Dengug antibody-free animals mor s.c. inoculated with 0.5 ml of either DEN-2(2.1 x 103 pfu/l; 3.8 x 106MIDSOAL), EM-4 (1.2 x 103 pfu/ml; 1.9 x 104 mso/)or a 1.l m.xtuxe of DW-2 and 4 vwcinrs, Approxinately sIx n ths after vaoci-nation, Mnls a ,re challenged with 104 MIDS0 (25 pfuLof wild type =1-2 virus,

DO ,J.,,. 1473 c.atnowor tNOV as is .,bo.aE59Ia111ITY CLAMFICAl0 OF T11s PAQE (;=am *e. 'M,..

20. Abstract (ont.)

It was found that:

i.) rM2-2/S-l animals slowed an anaimestic response as well as undetectableviremia after challenge with w.t. EEN-2 virus. Divalent vaccinees alsoshowed an ananmestic response after JDW-2 virus challenge.

(2.> EI-4 vaccine did not elicit a measurable immune response in 6/8 animalsas judged by PRNT, augmented N tests and by DE-2 challenge experiments.An anammuestic response was observed in 2/8 animals.

(3. N antibody levels of nost of the vaccinated animals declined markedlyon or before post vaccination day 150.

(4t Viral interference aung the two different vaccinesarotypes could notbe dmostrated.

Accession For

NTIS GRA&IDTIC TAB [1Unannounced 1Justification

By

Distribution/Availability Codes

Avail and/orist Special

Accouplishments: Dengue antibody-free animals were divided into four experi-

mental groups. Group I (8 monkeys) were s.c. inoculated with 0.5 ml of a 1:1

diluted W 2-2/S-l vaccine (2.9 x 103 pfu/ml; 3.8 x 106 MID5 0/ml). Group II

(8 monkeys) were inoculated with 0.5 ml of a 1:1 diluted IDW-4 vaccine

(1.2 x 103 pfu /ml, 1-9 x 104 MID 5 /ml). Group III (13 monkeys) were inoc-

ulated with a 1:1 mixture of DM-2 and DEN-4 vaccines. Group IV consisted of

uninoculated (control) monkeys. Experimental animal were bled on post

infection days 30, 60, 90, 120, 150, and 177 and N antibody titers against

MI~-2 and 4 vins were determined. nesults fmn these experiments are

shan in table 1. It can be seen that 6/8 DEN-2/S-1 vaccinated but only

2/8 r04-4 vaccinated monkeys showed antibody response against the homologous

wild type virus. 7/13 and 2/13 group III monkeys showed N antibodies against

hmmlogous wild type rR4-2 and M4-4 virus, respectively. A X2 test revealed

that there was no statistically significant difference in seroconversion

between movalent and divalent vaccinees (p-.5%). This dernonstrated that

viral interference among the two different vaccine serotypes did not occur.

In agreement with previous reports,(1) antibody levels of most of the vaccinated

animals declined markedly on or before post vacination day 150.

PR13s showed that the EM-4 vaccine did not elicit a strong immune response

in rhesus monkeys. This vas in agreement with results obtained in WRAIR whid

showed that only 2/5 human volunteers seroconverted after EN-4 Vaccination

(r, W. Brandt, personal omvunLcation).

Six months after vaccination, all animals were dallenged with 10,000

M 0 (25 pfui 1f000 monkey II)0) of EIN-2 (PR159) virus (see Table II, page

VI-8L Post challenge vireuia was detected in 3/4 group 4 (control) monkeys,

1/13 grou 3 (divalent vaccinees) and 5/8 group 2 (DEN-4 vaccimems). No

-1- -1

Vixunia was dtected in group 1 (MI-2 vaccinees).

As exected, all EEN-2 inoculated control animals uhwe a typical prin ry

antibody respns against mA-2 virus. mi-2/S-1 Vaccinees showed an anamestic

re~oe against th allenge virs. This type of antibody response was also

observed in 2/8 group 2 vaccinees and in all grou 3 vaccinees. Therere,

although some animals failed to show a measurable primary antibody response after

vaccination, challenge with w.t. rEN-2 showed that infection with vaccine did

ocur. We interpreted this data to mean that the standard PRT used (2) was not

the most sensitive assay to measure animal seroconversion. 7his hypothesis was

tested in our laboratory using the "augmented" neutralizing antibody test. Using

this test, we were able to show antibody response to homologous virus in sara

taken from group 1 monkeys B851 and B853 and from group III nvnkeys B810, B817,

B825, B806 and B804. Antibody titers ranged from 15 to 80 and wre le

in sure of the animals after post vaccination day 30. This test failed to re-

veal dengue 4 antibody titers in deNgu-4 vaccLnees which did not sero=vrt,

as neasured by the standard PR4 test.

In ccnclusion, these experiments showed that:

1. In agreement with previous work of Scot et al (1) rD-2/S-1 vaccinated

animals showed an anauestic response as well as undetectable viremia

after challenge with w.t. M2 virus. Divalent vaccinees also showed

an anmmustic respon m after IE-2 vlus challenge.

2. E-4 vaccine did not elicit a anawuAle Izme response. $v 6/8

animals as judged by PfP appnted N tests And by JZI.- challenge

-pe-Izuents. An anmmstc rezpose was orved in 2/8 anJ .ls,

3. In agreement With Previous reportsN antibody level of most of the.

vaocinated animls declined markedly on or before post a tikx day 150.

4. Viral 1t-ezuae ammg the t different vaccine usrotypes could rot

be dIn trated.

2 -

TABLE I i ANTIBODY RESPONSE OF RHESUS MONKEYS VACCINATED WITHWRAIR STRAINS OF DENGUE VIRUS.

ANTIBODY TITERS ON POST-VACCINATION DAY

' M Monkey Vaccine

Number Type 30 60 90 120 150 177

3840 D2 40 86 68 27 < 10 410B851 D2 <10 <10 410 <10 < 10 -10B873 D2 -10 72 14 £10 C10 <.10B. 3814 D2 .410 14 84 74 80 18

3837 D2 74 11 17 33 26 37B833 D2 40 160 80 150 159 320B853 D2 <10 <10 <10 < 10 < 10 <10B815 D2 410 32 29 •10 <.10 <10

B802 D4 -10 20 17 <10 <10 <10B829 D4 410 <•10 < 10 (10 <10 <10B812 D4 <.10 <10 <10 410 <10 < 10B805 D4 £10 410 <10 410 <10 <10

B826 D4 <10 <10 <10 <10 4 10 <10B834 D4 <10 <10 <10 410 < L0 <10B838 D4 52 27 <10 <10 < 10 <10B855 D4 <10 ,10 <10 <10 (10 <10

D2 D4 D2 D4 D2 D4 D2 D4 D2 D4 *D2

B811 D2+D4 18 <10 30 410 230 <10 110 <10 220 <10 96

B810 D2+D4 <10 <10 <10 <10 <10<10 <10<10 <-10 (10 <10

B813 D2+114 10 <10 72 410 310 <10 380 <10 270 (10 200B822 D2+D4 115 <10 >160 <10 >160 <10 >160 <10 >160 (10 $003827 D2+D4 58 <10>160 12 64>40 50 27 58 <10 130B817 D2+D4 <10 <10 (10 <0 <10410 <10(10 <10 <10 <103852 D2+D4 (10 <10 >640 >40 >640 29 >640 <10 590 (10 8803835 D2+D4 11 <10 72 <10 120 -10 230 <10 125 .10 220B825 D2+D4 <10 <10 <10 <10 <10<10 <10 (10 <10 <10 C103845 D2+D4 <10 <10 135 <10 47 <10 15 <10 13 <10 103836 D2+D4 <10 (10 <10 <10 <10<10 <10(410 <10 £10 <103806 D2+D4 <o10 <10 <o10 (1 <10<10 <oo10-C <10 <10 <103804 D2+D4 <10 <10 <10 <10 <I0 <10 <10 0O <10 <10 <10

3832 Control X contwl1 iAN

3831 Control3839 Control3286 Control <10 <10 <10 <10 4tO <10

1311 Control3521 ControlA962 Control' A1 Control

i ...... .; -....... ..- 3 -

10YADLI II

VUrea end Antibody Riaqionm of Vaccinated teus Monkeys After Challenge With Wild TypeDengue-2 Virus

Antibody titersYMkey Vaccine PRMT(Aug. TEst)* on Challenge day Day of viremia D-2 i titers on postumnber Type Seroconversuon (177 days) (pfu/nl) challenge days

D-2 D-4 D-2 D-4 10 30 605840 D2 + N.D. 110 N.D. - 86 v,640 8003851 D2 -(+) N.D. '10 N.D. - 380 540 6605873 D2 + N.D. d10 N.D. - 18 '640 20002814 D2 + N.D. 18 N.D. - 300 700 705837 )2 + N.D. 37 N.D.- - 500 -'640 960BS33 !2 + N.D. 320 N.D. - 490 >640 440M853 D2 -(+) N.D. '10 N.D; - >640 >640 1400B815 12 + M.D. 410 N.D. - 28 '640 N.D.*

38(2 4 M.D. + ,410 N.D. 6(7.5),7(5) A640 *640 3100B829 D4 N.D. -(-) -10 N.D. - '10 250 900B512 D4 N.D. -(-) 410 M.D. 5(2.5),7(2.5),8(7.5) -10 7640 1250JJ8$C 14 N.D. -(-) d10 N.D. - '10 350 6405826 D4 M.D. -(-) N10 M.D. - '10 >640 6205834 D4 N.D. -(-) -10 M.D. 6(2.5) 410 7320 >12801838 D4 M.D. + '10 N.D. 6(30),7(17.5) 225 >640 10501855 D4 N.D. -(-) 110 M.D. 7(10) '10 640 580

B811 D2+D4 + - 96 N.D. - 3000 7640 >25605810 D2+D4 -(+) -(-) N.10 M.D. - 320 7640 P25605813 D2+DI + - 200 N.D. - 190 135 1505822 D2+D4 + - 500 M.D. - 2800 7640 18005827 D2+D4 + + 130 M.D. - > 1280 7640 640B917 D2+D4 -(1) -() 10.D. 520 P640 72560B8.52 D2+I)4 + + 880 M.D. - 2000 ;P640 10005835 D2+D4 + - 220 N.D. - 75120 71640 6005825 D2+4 -(+) -(+ ) '1.0 N.D. - 245 7640 2805845 D2+D4 + - 10 N.D. - 7160 7640 2560!836 D2+De -(+) -(-) -'10 M.D. 5(2.5),6(7.5) 270 ;P640 2300B80P2+ -(+) -(-) '-10 M.D. - 620 ,640 72560i80 P2,.DIe -(4-) -(-) '-10 N.D. - 7640. '640 72560

5286 Control - - -10 410 8(2.5) '10 1300 125605311 Control - - £10 (.10 8(7.5) 410 640 1153521 Control - - e.10 4,10 6(10),7(25),8(5),9(2.5) '10 )640 #2560B11 Control - - '10 '10 '-10 Y640 >2560i832 Control - - '10 -10 - (Uninfected) '"10 '10 '105831 Control - - '10 -10 - (Uninfected) '10 '10 <105839 Control - - '-10 4'10 - (Uninfected) -10 '-10 410A62 Control - -. '-10 f-10 - (Uninfected). --10 '-10 C-10

*4.D.s eT oLoky died on pout-challenge day 51 of reasons unrelated to dengue virus infection.

mDs not do".

C+)s Kadlcates, wroonvermLon to homologous virus as determned by tlm augmented neutralizatlontests

-4-

2t-S

List of 1ferences

1. Scott, R MO, Nisalak, A., Eckels, K.H., Tingplapong, 14., Harrison, V.H.,Gould, D.J., Chapple, F.E., and Russell, P.K.: Dengue 2 vacine, viremiaand Inmune response in rhesus monkeys. Infect. Immun. 27, 181-186, 1980.

2. Russell, P.K., Nisalak, A., Sukhavachana, P., and Vivona, S.: A plaquereduction test for dengue virus neutralizing antibodies. J. Immnol.99, 285-790, 1976.

3. Sato, H., Albrecht, P., Krugman, S., and Ennis, F.: Sensitive neutraliza-tian test for Rubella antibody. J. Clin, Microbiol. 9, 259-264, 1979.

4. Asano, Y., Albrecht, P., Stagno, S. and Tacahashi, M. : Potentration ofneutralization of varicella-zoster virus by antibody to iumrnoglobulin.J. Infect. Dis. 146, 524-529, 1982.

5

In cadwtig the xeseAr scribed in this report, the investigator(s)

adhered to the "Qui for the Care and Use of Laboratory Animals,"

prepared by the Ccumuittee on Care and Use of Laboratory Animals of the

Institute of Laboratory Animal Resources, National Research Coucil

(DHEW Publication No. (NIH) 78-23, Revised 1978)

-6- 1iJ

6 A

AD-A143 082 EVALUATION IN RHESUS MONKEYS OF A BIVALENT L IVE *I /0ATTENUATED DENGUE VACCINE CONTAINING TYPES 2 AND 4VIRUSESIUI PUERTO RICO UNIV SAN JUAN E KRAISELBURD

INCAYSIFIED JUN 84 DAMO17 83-C-3029 F/G 6/5 NI

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INFOFThIATIONe

DEPARTMENT OF THE ARMY,•' US ARMY MIDICAL I!M(ARCH AND DEVELOPMENT COMMAND

FORT DETRICK FRIEIEICK MD 21701 5012

' RPLY TOATTENTION OF

SGRD-RHS 10 January 1985

Subject: Errata to Technical Report, ADA143082

AdministratorDefense Technical Information CenterATTN: DTIC-DDAC, Hrs. CrumbackerCameron StationAlexandria, VA 22304-6145

1. Reference:

a. Subject document, ADA143082, Final Report, Jun 84, for Contract No.DAMD1 7-83-C-3029.

b. Revised DD Form 1473 for subject document. (Incl 1)

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3. If you require further information or have questions regarding this matterplease contact Mrs. Jane Idoine of this office at Autovon 343-7253.

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REPORT DOCMENTATION PAGE DaI opLT4 a

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